CaMKII enhances voltage-gated sodium channel Nav1.6 activity and neuronal excitability

Abstract

<jats:p>Nav1.6 is the primary voltage-gated sodium channel isoform expressed in mature axon initial segments and nodes, making it critical for initiation and propagation of neuronal impulses. Thus, Nav1.6 modulation and dysfunction may have profound effects on input-output properties of neurons in normal and pathological conditions. Phosphorylation is a powerful and reversible mechanism regulating ion channel function. Because Nav1.6 and the multifunctional Ca<jats:sup>2+</jats:sup>/CaM-dependent protein kinase II (CaMKII) are independently linked to excitability disorders, we sought to investigate modulation of Nav1.6 function by CaMKII signaling. We show that inhibition of CaMKII, a Ser/Thr protein kinase associated with excitability, synaptic plasticity, and excitability disorders, with the CaMKII-specific peptide inhibitor CN21 reduces transient and persistent currents in Nav1.6-expressing Purkinje neurons by 87%. Using whole-cell voltage clamp of Nav1.6, we show that CaMKII inhibition in ND7/23 and HEK293 cells significantly reduces transient and persistent currents by 72% and produces a 5.8 mV depolarizing shift in the voltage-dependence of activation. Immobilized peptide arrays and nanoflow LC-ESI/MS of Nav1.6 reveals potential sites of CaMKII phosphorylation, specifically Ser561 and Ser641/Thr642 within the first intracellular loop of the channel. Using site-directed mutagenesis to test multiple potential sites of phosphorylation, we show that Ala substitutions of Ser561 and Ser641/Thr642 recapitulate the depolarizing shift in activation and reduction in current density. Computational simulations to model effects of CaMKII inhibition on Nav1.6 function demonstrate dramatic reductions in spontaneous and evoked action potentials in a Purkinje cell model, suggesting that CaMKII modulation of Nav1.6 may be a powerful mechanism to regulate neuronal excitability.</jats:p>

T-type Ca2+and persistent Na+currents synergistically elevate ventral, not dorsal, entorhinal cortical stellate cell excitability

“…The possibility, though, that Ca 2+ entry via T-type Ca 2+ channels may also rapidly and reversibly phosphorylate Na + channels to enhance the persistent Na + current in ventral mEC layer II stellate neurons cannot be excluded. Certainly Ca 2+ -dependent modification of Na V 1.6 channels, which principally encode for persistent Na + currents in neurons (Hargus et al, 2013; Katz et al, 2018), increased the persistent Na + current in hippocampal CA1 pyramidal neuronal dendrites (Yu et al, 2018) and cerebellar Purkinje neurons (Zybura et al, 2020). Moreover, Ca 2+ - calmodulin dependent kinase, CaMKII, phosphorylates Na V 1.2 channels, that predominantly underlie transient Na + currents, to generate a persistent Na + current in heterologous systems (Thompson et al, 2017).…”

The Calcium/Calmodulin-Dependent Kinases II and IV as Therapeutic Targets in Neurodegenerative and Neuropsychiatric Disorders

“…Moreover, the aberrant phosphorylation of CaMKII is the cause of neurological symptoms of Angelman syndrome [ 39 ]. However, since CaMKII is expressed most abundantly in neurons, it regulates many aspects of neuronal function, including neurotransmitter synthesis and release, modulation of ion channel activity, cellular transport, cellular morphology, and neurite extension [ 76 , 77 , 78 , 79 ].…”

“…Studies showed that this kinase regulates the conductivity of the various types of sodium, potassium, and calcium channels [ 84 ]. Considering the neuronal cells, CaMKII enhances voltage-gated sodium channel Nav1.6 activity and neuronal excitability [ 79 ]. Moreover, by modulating the voltage-gated calcium (CaV)2.1 channels that conduct P/Q type Ca 2+ currents, CaMKII regulates the neurotransmitter release [ 85 ].…”

CaMKII in Regulation of Cell Death During Myocardial Reperfusion Injury

“…Since the first CaMKII inhibitor, KN-62 was invented in experiment (Tokumitsu et al, 1990); more and more drugs targeting CaMKII were discovered. Yet most of drugs are still utilized in animal models, only three specific CaMKII inhibitors, rimacalib (SMP-114), tatCN21 (Zybura et al, 2020), and ranolazine are applied in clinical trials, so there is still a long journey to the development of CaMKII-related medicine and its utilization in myocardial I/R injury. Meanwhile, many commonly used drugs have effect on CaMKII suppression, which is also unfavorable to the passion of targeting CaMKII invention.…”